Proposal Summary Elucidating the fundamental mechanisms of enzyme regulation is paramount for disease prevention and treatment. Aberrant enzyme activity is a chief factor in the manifestation of diseases including rheumatoid arthritis, neurodegenerative disorders, and cancer. Specifically, dysregulation of catalytically active ADAM (a disintegrin and metalloprotease) protein family members directly contributes to excessive TNF-? release in rheumatoid arthritis, processing of amyloid precursor protein (APP) in Alzheimer's Disease, and augmented growth factor and cytokine release in tumor establishment and metastatic spread. ADAMs are ectodomain sheddases that transform latent cell-bound substrates to soluble, biologically active derivatives. However, a major obstacle in understanding ADAM regulation/dysregulation in normal and disease states is that of the 21 human ADAMs, nearly half lack the active site consensus sequence indicative of a functional metalloprotease.. Preliminary data from our lab provide evidence for a novel regulatory mechanism for ADAM sheddase activity involving noncatalytic members. The current study posits that noncatalytic ADAMs ?mimic? select aspects of catalytic ADAMs t0 compete for molecular binding partners (e.g. integrin adhesion receptors) essential for catalysis. This novel regulatory model, tentatively termed competitive mimicry?, embodies an unexplored aspect of enzyme regulation with potential broad implications, as 92 of the superset of ~570 human proteases (metallo, serine, cysteine, etc.) inexplicably lack catalytic consensus elements. In Aim 1, we will determine how catalytic and noncatalytic ADAM members are biochemically, structurally, and evolutionarily related. Bioinformatics, kinetics, purified enzymatic assays, and crystallographic approaches will be used to delineate the first functional link between noncatalytic and catalytically active ADAMs. In Aim 2, we will determine how noncatalytic ADAMs govern the shedding activity of catalytically active ADAM counterparts. Using mutagenesis and protein engineering approaches, this objective will define the regions and protein domains of noncatalytic ADAMs required for the newly identified regulatory function. This proposal is innovative as it is a significant departure from established mechanisms of enzyme regulation through studying a neglected group of rather abundant ?dead? enzymes. The novelty of the work is that the premise of ADAM competition allows for an unexplored, but testable, aspect of ADAM biology that accounts for the number of noncatalytic members. The proposal is significant because it will: i) establish the first functional role of noncatalytic ADAMs in catalytic regulation, ii) yield novel contexts and targets for therapeutic development in diseases associated with enzyme dysregulation, iii) foster and enhance a research culture at a new medical school in a historically underserved region and iv) provide training to undergraduate and graduate students in a variety of molecular, biochemical, bioinformatic, and cellular techniques, consistent with the mission of an R15 application.